The bronopol and kathon are chemical preservative which prevent degradation of milk samples and maintain authenticity in analysis. The detection is based on HPLC-UV-Vis spectroscopy, in which C18 column (250 mm × 4.6 mm, 5 µm) was used for chromatographic separations, with a mobile phase comprising 0.1% phosphoric acid in water: Methanol: 0.1% phosphoric acid in acetonitrile (80:10:10) at a flow rate 0.8 ml/min at ambient temperature and with the UV detection at 250 nm for bronopol and 274 nm for kathon. The retention time of bronopol, kathon (MI 2-methyl-4-isothiazolin-3-one) and kathon (CMI 5-chloro-2-methyl-4-isothiazolin-3-one) was 4.52 min, 3.98 min and 6.68 min respectively with a total run time of 10 min. The linearity of the method was satisfactory with regression coefficient (R2) = 0.99. The limit of quantification was 72, 240, 390 mg L-1 for bronopol, kathon (MI) and kathon (CMI) respectively. Five spiked levels (62.5, 125, 250, 500 and 1000 mg L-1) were used to determine the recovery of bronopol, kathon (MI) and kathon (CMI) which was found to be 95.41 ± 11.84, 95.75 ± 8.21 and 92.22 ± 14.64% respectively, with relative standard deviations in the range 5.9-14.6%. The standardized analytical method was successfully used to rapidly detect bronopol and kathon in milk samples.
Keywords: Bronopol; CMI; HPLC; Kathon; MI.